1. Field of the Invention
The present invention relates to a process for producing sporangia of microorganisms belonging to Bacillus popilliae containing spores and parasporal bodies use as control agents of Scarabaeidae insects (“sporangia containing spores and parasporal bodies” may simply be referred to as “sporangia”).
2. Description of the Related Art
The larva of Scarabaeidae insects feed on a wide range of plant roots such as those of grasses, agricultural and horticultural crops and trees, and are known to cause considerable damage. Since these larva live underground, in addition to it being difficult to identify the locations where these larva are present, a high degree of control effects are unable to be obtained even if agricultural chemicals are sprayed from the air. Therefore, in order to demonstrate remarkable effects, it has been necessary to spray large amounts of agricultural chemicals over a wide range to enable the chemicals to penetrate into the ground. However, since there are concerns over this method having a detrimental effect on both the natural environment and people, a more effective control method is desired.
Microorganisms belonging to Bacillus popilliae are known to parasitically cause milky disease in the larva of Scarabaeidae insects, eventually cause their death. Attempts have long been made to use the sporangia of these microorganisms to control Scarabaeidae insects on which agricultural chemicals have little effect. However, although these microorganisms grow within the larva of Scarabaeidae insects, it is difficult to grow them by culturing using artificial media, and the production of sporangia of these microorganisms in media has been particularly difficult. In addition, Fukuhara reported that infection and pathogenesis of larva do not occur with sporangia obtained by culturing using conventional media (Fukuhara, T., ed., Insect Pathology, p. 57, 1979). Namely, there has yet to be an efficient method for large-volume production of these sporangia that can be used effectively for control of Scarabaeidae insects.
For example, Haynes, et al. reported an example of obtaining a maximum of 2.06×107 sporangia per 1 ml of liquid culture by attempting to culture microorganisms belonging to Bacillus popilliae in liquid medium containing 0.5% peptone, 1.5% yeast extract, 0.3% dipotassium hydrogenphosphate, 0.1% glucose and 1% activated carbon (Journal of Invertebrate Pathology, Vol. 22, p. 377-381, 1973). However, the ratio of the content of glutamic acid to the medium and the ratio of the content of glutamic acid to total amino acids are unclear. In addition, Haynes, et al. also described in the same report that amino acid composition is unrelated to production of sporangia (p. 379, column 1, line 19).
In addition, Haynes, et al. reported that 3.1×107 sporangia per 1 ml of liquid culture were obtained by culturing mature cells of Bacillus popilliae in the late logarithmic increase stage in liquid medium containing 0.5% peptone (tryptone), 1.5% yeast extract, 0.3% dipotassium hydrogenphosphate, 0.1% glucose and 1% activated carbon (Journal of Invertebrate Pathology, Vol. 19, p. 125-130, 1972). However, this culturing method has a long culturing time, taking roughly two weeks.
In addition, an example of having obtained 1×109 spores per 1 ml of liquid culture by culturing in liquid medium containing 1% soluble starch, 0.1% trehalose, 0.5% yeast extract, 0.3% dipotassium hydrogenphosphate and 0.1% calcium carbonate is indicated in the specification of U.S. Pat. No. 4,824,671. However, in this case as well, the resulting spores were not accompanied by parasporal bodies, and the rate of infection with milky disease when spores were sprayed at the rate of 2.0×1012 spores per 1 kg of soil and allowed to be orally ingested by larva of Scarabaeidae insects for 7 weeks was 47.50%, indicating weak insecticidal effects on larva of Scarabaeidae insects even when compared with sporangia formed within the bodies of the larva.